The trace element copper (Cu) is an essential cofactor for critical life processes that include mitochondrial oxidative phosphorylation, iron absorption, reactive oxygen detoxification, peptide hormone biogenesis, connective tissue maturation and cellular signaling. Dietary Cu limitation, defects in the Cu acquisition machinery, or the inability to adapt to Cu deficiency gives rise to pathophysiological states in specific cells and tissues including iron deficiency anemia, neuronal and cognitive dysfunction, connective tissue defects, hypertrophic cardiomyopathy and gestational lethality. Importantly, Cu chelators are used to treat patients with devastating Cu overload disease, and are being developed for use in cancer chemotherapy. While Cu acquisition, and appropriate adaptive responses to Cu limitation are critical for human development and health, we know little about how cells adapt to Cu limitation through increased Cu import or critical regulatory changes. This application details experiments to decipher two fundamentally important gaps in our knowledge of the components involved in Cu acquisition and the mechanisms by which cells adapt to Cu-limitation. We will exploit the powerful fungal model system, Cryptococcus neoformans, which has a strong reliance on Cu- dependent metabolism for its growth, and which has a rich biology of genes whose expression is regulated by Cu-deficiency. First, we will elucidate the role that a new extracellular protein, Bim1, plays in Cu acquisition. We will test the hypothesis that Bim1 serves as an extracellular Cu ligand that functions in concert with the Ctr1-mediated plasma membrane Cu+ import machinery and that Bim1 is critical for cell growth and survival in a naturally occurring Cu-limiting environment. These experiments will validate the first extracellular Cu-ligand in eukaryotic Cu-uptake, filling a gap in our understanding of how organisms acquire this essential trace element. Second, we will decipher how under Cu-limiting conditions, expression of the abundant Sod1 Cu-dependent protein is extinguished, the physiological role this plays in prioritizing Cu to proteins critical for normal growth, and how cells adapt to loss of Sod1 through changes in the localization and synthesis of other proteins via cellular translation reprogramming. Together, these studies will advance new concepts for how cells acquire the essential metal ion Cu, invoke adaptive responses to Cu limitation and regulate these processes for normal cell growth, function and health.